Warfarin is a commonly used anticoagulant. Taken during pregnancy, it causes malformations and fetal death in humans. Study of warfarin's teratogenesis has been hindered by the lack of an animal model suitable for morphogenic and biochemical studies. Using embryonic mouse forelimb cultures, I have developed an in vitro model of warfarin's disruption of chondrogenesis. This proposal is designed to investigate the molecular and developmental basis for this disruption. Chondrogenesis in the forelimb involves morphologically distinct events and structures which can be used to evaluate warfarin's effect. Histological studies will be done to investigate warfarin's effect on mesenchymal condensation and the morphology of the apical ectodermal ridge. If mesenchymal condensation is abnormal study of mesenchymal cell adhesion will be done. A study by myself and Holmes shows that warfarin causes a lasting depression of 3H-thymidine incorporation in cultured forelimbs. Two of the possible causes of this decrease are cell death which would decrease the number of cells incorporating 3H-thymidine and cytodifferentiation which is associated with a decreased mitotic activity. The possibility that cessation of proliferation may cause premature cytodifferentiation, will be studied by evaluating the effect of a known mitotic inhibitor on chondrogenesis. Cell death is of interest because it has been proposed as a mechanism of teratogenesis. Warfarin has been shown to inhibit the synthesis of gamma carboxyglutamic acid (GLA). GLA will be assayed for in forelimbs and if found, warfarin's effect on its synthesis will be studied. Since GLA chelates Ca ions, the ion's effect on chondrogenesis will be studied.